Plant regeneration from mesophyll protoplasts of Dianthus superbus

Plant regeneration from mesophyll protoplasts of Dianthus superbus

Leaf mesophyll protoplasts of Dianthus superbus were cultured at a density of 5x10⁴protoplasts/ml and divided at about 18% plating efficiency in MS liquid medium supplemented with 0.5 mg/L BAP, 2.0 mg/L NAA and 9% mannitol after 2 weeks. Protocolonies formed after 3 to 4 weeks of culture in the dark at 27℃. These colonies were transferred to continuous illumination (21.5Em-² sec-¹) for 2 weeks where most of the colonies divided to form microcalli, about 2 mm in diameter. Subsequently, green microcalli were transferred to MS solidified medium with 2.0 mg/L 2,4-0 that induced shoot- forming calli after 4 weeks. These calli were transferred onto N,-2 medium containing 0.1 mg/L 2,4-0, 0.1 mg/L NAA, 2.0 mg/L kinetin and 2.0 g/L casein hydrolysate and were cultured under light. After 5 weeks the calli gave rise to multiple shoots 00 to 15 per callus). Upon transfer to MS medium containing 2.0 mg/L NAA, individual shoots were rooted in 4 weeks. The regenerants were successfully transplanted into potting soil